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Journal of Bacteriology, January 2000, p. 135-142, Vol. 182, No. 1
0021-9193/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Identification of a New Class of 5'-Adenylylsulfate (APS) Reductases from Sulfate-Assimilating Bacteria

Julie Ann Bick, Jonathan J. Dennis, Gerben J. Zylstra, Jason Nowack, and Thomas Leustek*

Biotechnology Center for Agriculture and the Environment, Rutgers University, New Brunswick, New Jersey 08901-8520

Received 23 July 1999/Accepted 11 October 1999

A gene was cloned from Burkholderia cepacia DBO1 that is homologous with Escherichia coli cysH encoding 3'-phosphoadenylylsulfate (PAPS) reductase. The B. cepacia gene is the most recent addition to a growing list of cysH homologs from a diverse group of sulfate-assimilating bacteria whose products show greater homology to plant 5'-adenylylsulfate (APS) reductase than they do to E. coli CysH. The evidence reported here shows that the cysH from one of the species, Pseudomonas aeruginosa, encodes APS reductase. It is able to complement an E. coli cysH mutant and a cysC mutant, indicating that the enzyme is able to bypass PAPS, synthesized by the cysC product. Insertional knockout mutation of P. aeruginosa cysH produced cysteine auxotrophy, indicating its role in sulfate assimilation. Purified P. aeruginosa CysH expressed as a His-tagged recombinant protein is able to reduce APS, but not PAPS. The enzyme has a specific activity of 5.8 µmol · min-1 · mg of protein-1 at pH 8.5 and 30°C with thioredoxin supplied as an electron donor. APS reductase activity was detected in several bacterial species from which the novel type of cysH has been cloned, indicating that this enzyme may be widespread. Although an APS reductase from dissimilatory sulfate-reducing bacteria is known, it shows no structural or sequence homology with the assimilatory-type APS reductase reported here. The results suggest that the dissimilatory and assimilatory APS reductases evolved convergently.

* Corresponding author. Mailing address: Biotech Center, Rutgers University, 59 Dudley Rd., New Brunswick, NJ 08901-8520. Phone: (732) 932-8165, ext. 326. Fax: (732) 932-0312. E-mail: LEUSTEK{at}AESOP.RUTGERS.EDU.

Journal of Bacteriology, January 2000, p. 135-142, Vol. 182, No. 1
0021-9193/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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